Distinct functions of ATG16L1 isoforms in membrane binding and LC3B lipidation in autophagy-related processes

• Published in: Nature cell biology Vol. 21; no. 3; pp. 372 - 383
• Main Authors: Lystad, Alf Håkon, Carlsson, Sven R., de la Ballina, Laura R., Kauffman, Karlina J., Nag, Shanta, Yoshimori, Tamotsu, Melia, Thomas J., Simonsen, Anne
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01.03.2019; Nature Publishing Group
• Subjects: 13; 13/109; 14; 14/19; 14/35; 38; 38/1; 38/35; 38/70; 631/80/313; 631/80/39; 631/80/39/2346; 631/92/287; 82; 82/1; 82/16; 82/29; 82/80; 82/83; Amino Acid Sequence; Amino acids; Animals; Autophagy; Autophagy (Cytology); Autophagy-Related Proteins - genetics; Autophagy-Related Proteins - metabolism; Binding; Binding Sites - genetics; Biomedical and Life Sciences; C-Terminus; Cancer Research; Cell Biology; Cell Membrane - metabolism; Depletion; Developmental Biology; Endosomes; Endosomes - metabolism; GABARAP protein; HEK293 Cells; Humans; Isoforms; Life Sciences; Medical schools; Membrane Lipids - metabolism; Membrane structures; Membranes; Mice; Microtubule-Associated Proteins - metabolism; Phagocytosis; Phosphatidylethanolamine; Phospholipids; Post-translational modifications; Protein Binding; Protein Isoforms - genetics; Protein Isoforms - metabolism; Protein research; Proteins; RAW 264.7 Cells; Regulatory genes; Sequence Homology, Amino Acid; Stem Cells
• ISSN: 1465-7392; 1476-4679; 1476-4679
• DOI: 10.1038/s41556-019-0274-9

Covalent modification of LC3 and GABARAP proteins to phosphatidylethanolamine in the double-membrane phagophore is a key event in the early phase of macroautophagy, but can also occur on single-membrane structures. In both cases this involves transfer of LC3/GABARAP from ATG3 to phosphatidylethanolamine at the target membrane. Here we have purified the full-length human ATG12-5–ATG16L1 complex and show its essential role in LC3B/GABARAP lipidation in vitro. We have identified two functionally distinct membrane-binding regions in ATG16L1. An N-terminal membrane-binding amphipathic helix is required for LC3B lipidation under all conditions tested. By contrast, the C-terminal membrane-binding region is dispensable for canonical autophagy but essential for VPS34-independent LC3B lipidation at perturbed endosomes. We further show that the ATG16L1 C-terminus can compensate for WIPI2 depletion to sustain lipidation during starvation. This C-terminal membrane-binding region is present only in the β-isoform of ATG16L1, showing that ATG16L1 isoforms mechanistically distinguish between different LC3B lipidation mechanisms under different cellular conditions. Lystad et al. identify distinct membrane binding regions in ATG16L1 and show that the β-isoform-specific C-terminal region is required for VPS34/ULK1/2-independent non-canonical autophagy.